Angiopoietin-like protein 2 promotes chondrogenic differentiation during bone growth as a cartilage matrix factor.

OBJECTIVE: Chondrocyte differentiation is crucial for long bone growth. Many cartilage extracellular matrix (ECM) proteins reportedly contribute to chondrocyte differentiation, indicating that mechanisms underlying chondrocyte differentiation are likely more complex than previously appreciated. Angiopoietin-like protein 2 (ANGPTL2) is a secreted factor normally abundantly produced in mesenchymal lineage cells such as adipocytes and fibroblasts, but its loss contributes to the pathogenesis of lifestyle- or aging-related diseases. However, the function of ANGPTL2 in chondrocytes, which are also differentiated from mesenchymal stem cells, remains unclear. Here, we investigate whether ANGPTL2 is expressed in or functions in chondrocytes. METHODS: First, we evaluated Angptl2 expression during chondrocyte differentiation using chondrogenic ATDC5 cells and wild-type epiphyseal cartilage of newborn mice. We next assessed ANGPTL2 function in chondrogenic differentiation and associated signaling using Angptl2 knockdown ATDC5 cells and Angptl2 knockout mice. RESULTS: ANGPTL2 is expressed in chondrocytes, particularly those located in resting and proliferative zones, and accumulates in ECM surrounding chondrocytes. Interestingly, long bone growth was retarded in Angptl2 knockout mice from neonatal to adult stages via attenuation of chondrocyte differentiation. Both in vivo and in vitro experiments show that changes in ANGPTL2 expression can also alter p38 mitogen-activated protein kinase (MAPK) activity mediated by integrin α5ß1. CONCLUSION: ANGPTL2 contributes to chondrocyte differentiation and subsequent endochondral ossification through α5ß1 integrin and p38 MAPK signaling during bone growth. Our findings provide insight into molecular mechanisms governing communication between chondrocytes and surrounding ECM components in bone growth activities.

A physical map with yeast artificial chromosome (YAC) clones covering 63% of the 12 rice chromosomes.

A new YAC (yeast artificial chromosome) physical map of the 12 rice chromosomes was constructed utilizing the latest molecular linkage map. The 1439 DNA markers on the rice genetic map selected a total of 1892 YACs from a YAC library. A total of 675 distinct YACs were assigned to specific chromosomal locations. In all chromosomes, 297 YAC contigs and 142 YAC islands were formed. The total physical length of these contigs and islands was estimated to 270 Mb which corresponds to approximately 63% of the entire rice genome (430 Mb). Because the physical length of each YAC contig has been measured, we could then estimate the physical distance between genetic markers more precisely than previously. In the course of constructing the new physical map, the DNA markers mapped at 0.0-cM intervals were ordered accurately and the presence of potentially duplicated regions among the chromosomes was detected. The physical map combined with the genetic map will form the basis for elucidation of the rice genome structure, map-based cloning of agronomically important genes, and genome sequencing.

Monosaccharide composition of sweetpotato fiber and cell wall polysaccharides from sweetpotato, cassava, and potato analyzed by the high-performance anion exchange chromatography with pulsed amperometric detection method.

The cell wall materials (CWMs) from sweetpotato (Ipomoea batatas cv. Kokei 14), cassava (Manihot esculenta), and potato (Solanum tuberosum cv. Danshaku) and commercial sweetpotato fiber as well as their polysaccharide fractions were analyzed for sugar composition by the high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) method. The separation of arabinose and rhamnose, and xylose and mannose, by this method has been improved using a CarboPac PA 10 column. Pretreatment of the CWMs and cellulose fractions with 12 M H(2)SO(4) was required for complete hydrolysis to occur. Commercial sweetpotato fiber was found to be mainly composed of glucose (88.4%), but small amounts of other sugars were also detected. Among the root crops, sweetpotato CWM had the highest amount of pectin and galacturonic acid. Fucose was detected only in cassava CWM and its hemicellulose fraction, while galactose was present in the highest amount in potato CWM. Among the polysaccharide fractions, it was only in the hemicellulose fraction where significant differences in the sugar composition, especially in the galactose content, were observed among the root crops.

Physical mapping of duplicated genomic regions of two chromosome ends in rice.

Two genomic regions duplicated in distal ends of the short arms of chromosomes 11 and 12 in rice (Oryza sativa L.) were characterized by YAC ordering with 46 genetic markers. Physical maps covering most of the duplicated regions were generated. Thirty-five markers, including 21 rice cDNA clones, showed the duplicated loci arrayed strictly in the same order along the two specific genomic regions. Regardless of their different genetic distances, the two duplicated segments may have a similar and minimum physical size with an expected length of about 2.5 Mb. However, differences of RFLP frequency for the duplicated DNA copies and recombination frequency for a given homoeologous area between the two regions were observed, indicating that these changes in genome organization occurred after the duplication. Our results establish a good model system for resolving the relationships between gene duplication, expression of duplicated genes, and the frequency of meiotic recombination in small chromosomal regions.

Physical mapping of the rice genome with YAC clones.

Construction of a rice physical map covered by YAC clones which have been arranged over half of the genome length is presented here. A total of 1285 RFLP and RAPD markers almost evenly distributed on the rice genetic map could select 2974 YAC clones and 2443 clones of them were located on their original positions. Rice YACs carrying 350 kb average insert fragments of 2443 clones could cover 222 megabase length of the rice genome, corresponding to 52% of the whole genome size (4.3 Mb). Chromosome landing with many YAC clones on the high-density genetic map loci efficiently integrated the genetic map with a physical map. This is the first step to generate a comprehensive genome map of rice. An integrated genome map should be an indispensable tool to figure out genome structure as well as to clone trait genes by map-based cloning.

Ordered YAC clone contigs assigned to rice chromosomes 3 and 11.

Yeast artificial chromosome (YAC) clones were arranged on the positions of restriction fragment length polymorphism (RFLP) and sequence-tagged site (STS) markers already mapped on the high-resolution genetic maps of rice chromosomes 3 and 11. From a total of 416 and 242 YAC clones selected by colony/Southern hybridization and polymerase chain reaction (PCR) analysis, 238 and 135 YAC clones were located on chromosomes 3 and 11, respectively. For chromosomes 3 and 11, 24 YAC contigs and islands with total coverage of about 46% and 12 contigs and islands with coverage of about 40%, respectively, were assigned. Although many DNA fragments of multiple copy marker sequences could not be mapped to their original locations on the genetic map by Southern hybridization because of a lack of RFLP, the physical mapping of YAC clones could often assign specific locations of such multiple copy sequences on the genome. The information provided here on contig formation and similar sequence distribution revealed by ordering YAC clones will help to unravel the genome organization of rice as well as being useful in isolation of genes by map-based cloning.

An ordered yeast artificial chromosome library covering over half of rice chromosome 6.

Yeast artificial chromosome (YAC) clones carrying DNA marker sequences located on the rice genetic map of chromosome 6 were ordered for physical mapping. A total of 122 restriction fragment length polymorphism markers, 16 sequence-tagged site markers, and five random amplified polymorphic DNA markers located, on average, at 0.9-cM intervals, were used for YAC clone screening by colony/Southern hybridization and PCR screening, respectively. A total of 216 individual YACs were selected from our YAC library of 7000 clones covering six genome equivalents. Each DNA marker could select, on average, 4.8 YAC clones, with 11 clones being the maximum. The YACs localized to the corresponding linkage map positions form 43 contigs and encompass about 60% of rice chromosome 6. This is the first step in constructing a physical map covering the whole rice genome by chromosome landing with YAC clones. These YACs and data will be used soon to isolate phenotypical trait genes by map-based cloning.

Regional distribution of 14C-zonisamide in rat brain.

Zonisamide (1,2-benzisoxazole-3-methane sulfonamide) is a new antiepileptic drug developed in Japan. This compound was proven to possess a strong inhibitory effect on convulsions of cortical origin, whether induced by electric or chemical stimuli. Regional distribution of 14C-zonisamide was investigated in rat brain using autoradiography. A high uptake of 14C activity was observed in the cerebral cortex and the midbrain. A pair-match analysis of primary motor cortex versus primary sensory cortex revealed a slightly higher uptake in primary motor cortex. In the cerebellum, a higher uptake was observed in the cortex than medulla. Sagittal section analyses revealed that a high uptake of 14C activity was observed in the cerebral cortex and colliculus, and a moderate uptake was seen in the cerebellum, thalamus, hypothalamus, and striatal body, thus suggesting the distribution of 14C-zonisamide is similar to that of flunitrazepam and phenytoin.

Mature teratoma arising from intra-abdominal contralateral undescended testis in an infant: a case report.

A 4-month-old male infant was seen because of an asymptomatic undescended left testis and a right sided abdominal mass. CT revealed a calcified retroperitoneal tumour. Histological examination of surgical specimens showed a mature primary teratoma of the contralateral undescended testis. While this is very rare, infants with undescended testis should be carefully examined to rule out intra-abdominal malignant tumours.